EP1856111B1 - Purification of mupirocin - Google Patents
Purification of mupirocin Download PDFInfo
- Publication number
- EP1856111B1 EP1856111B1 EP06723096.1A EP06723096A EP1856111B1 EP 1856111 B1 EP1856111 B1 EP 1856111B1 EP 06723096 A EP06723096 A EP 06723096A EP 1856111 B1 EP1856111 B1 EP 1856111B1
- Authority
- EP
- European Patent Office
- Prior art keywords
- mupirocin
- acid
- solution
- precipitate
- recovered
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- MINDHVHHQZYEEK-UHFFFAOYSA-N (E)-(2S,3R,4R,5S)-5-[(2S,3S,4S,5S)-2,3-epoxy-5-hydroxy-4-methylhexyl]tetrahydro-3,4-dihydroxy-(beta)-methyl-2H-pyran-2-crotonic acid ester with 9-hydroxynonanoic acid Natural products CC(O)C(C)C1OC1CC1C(O)C(O)C(CC(C)=CC(=O)OCCCCCCCCC(O)=O)OC1 MINDHVHHQZYEEK-UHFFFAOYSA-N 0.000 title claims abstract description 77
- 229960003128 mupirocin Drugs 0.000 title claims abstract description 71
- DDHVILIIHBIMQU-YJGQQKNPSA-L mupirocin calcium hydrate Chemical compound O.O.[Ca+2].C[C@H](O)[C@H](C)[C@@H]1O[C@H]1C[C@@H]1[C@@H](O)[C@@H](O)[C@H](C\C(C)=C\C(=O)OCCCCCCCCC([O-])=O)OC1.C[C@H](O)[C@H](C)[C@@H]1O[C@H]1C[C@@H]1[C@@H](O)[C@@H](O)[C@H](C\C(C)=C\C(=O)OCCCCCCCCC([O-])=O)OC1 DDHVILIIHBIMQU-YJGQQKNPSA-L 0.000 title claims abstract description 68
- 229930187697 mupirocin Natural products 0.000 title claims abstract description 66
- 238000000746 purification Methods 0.000 title abstract description 6
- 238000000034 method Methods 0.000 claims abstract description 32
- 239000002244 precipitate Substances 0.000 claims abstract description 17
- 239000003960 organic solvent Substances 0.000 claims abstract description 13
- 238000001556 precipitation Methods 0.000 claims abstract description 12
- 239000007864 aqueous solution Substances 0.000 claims abstract description 6
- 239000000243 solution Substances 0.000 claims description 32
- NTIZESTWPVYFNL-UHFFFAOYSA-N Methyl isobutyl ketone Chemical compound CC(C)CC(C)=O NTIZESTWPVYFNL-UHFFFAOYSA-N 0.000 claims description 18
- UIHCLUNTQKBZGK-UHFFFAOYSA-N Methyl isobutyl ketone Natural products CCC(C)C(C)=O UIHCLUNTQKBZGK-UHFFFAOYSA-N 0.000 claims description 18
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 15
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 15
- 238000000855 fermentation Methods 0.000 claims description 12
- 230000004151 fermentation Effects 0.000 claims description 12
- 239000002904 solvent Substances 0.000 claims description 12
- 238000005119 centrifugation Methods 0.000 claims description 10
- 238000002425 crystallisation Methods 0.000 claims description 7
- 230000008025 crystallization Effects 0.000 claims description 7
- 239000000203 mixture Substances 0.000 claims description 7
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 6
- 239000002253 acid Substances 0.000 claims description 6
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims description 6
- 238000001914 filtration Methods 0.000 claims description 6
- 238000001704 evaporation Methods 0.000 claims description 5
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 4
- 230000008020 evaporation Effects 0.000 claims description 4
- 238000004042 decolorization Methods 0.000 claims description 3
- 238000001035 drying Methods 0.000 claims description 3
- 150000002148 esters Chemical class 0.000 claims description 3
- 238000000605 extraction Methods 0.000 claims description 3
- 238000005352 clarification Methods 0.000 claims description 2
- 150000002576 ketones Chemical group 0.000 claims description 2
- MINDHVHHQZYEEK-HBBNESRFSA-N mupirocin Chemical compound C[C@H](O)[C@H](C)[C@@H]1O[C@H]1C[C@@H]1[C@@H](O)[C@@H](O)[C@H](C\C(C)=C\C(=O)OCCCCCCCCC(O)=O)OC1 MINDHVHHQZYEEK-HBBNESRFSA-N 0.000 abstract description 11
- 230000003115 biocidal effect Effects 0.000 abstract description 7
- 235000010633 broth Nutrition 0.000 description 18
- 238000003756 stirring Methods 0.000 description 14
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 12
- IMNFDUFMRHMDMM-UHFFFAOYSA-N N-Heptane Chemical compound CCCCCCC IMNFDUFMRHMDMM-UHFFFAOYSA-N 0.000 description 10
- 239000000284 extract Substances 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 6
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 6
- 239000000706 filtrate Substances 0.000 description 6
- 238000002360 preparation method Methods 0.000 description 6
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 5
- 229930194369 pseudomonic acid Natural products 0.000 description 5
- 239000003242 anti bacterial agent Substances 0.000 description 4
- 229940088710 antibiotic agent Drugs 0.000 description 4
- OCKGFTQIICXDQW-ZEQRLZLVSA-N 5-[(1r)-1-hydroxy-2-[4-[(2r)-2-hydroxy-2-(4-methyl-1-oxo-3h-2-benzofuran-5-yl)ethyl]piperazin-1-yl]ethyl]-4-methyl-3h-2-benzofuran-1-one Chemical compound C1=C2C(=O)OCC2=C(C)C([C@@H](O)CN2CCN(CC2)C[C@H](O)C2=CC=C3C(=O)OCC3=C2C)=C1 OCKGFTQIICXDQW-ZEQRLZLVSA-N 0.000 description 3
- 125000001931 aliphatic group Chemical group 0.000 description 3
- 239000002178 crystalline material Substances 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 239000012467 final product Substances 0.000 description 3
- 238000002955 isolation Methods 0.000 description 3
- 239000012074 organic phase Substances 0.000 description 3
- 238000011084 recovery Methods 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 description 2
- 241000589540 Pseudomonas fluorescens Species 0.000 description 2
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 2
- 125000000217 alkyl group Chemical group 0.000 description 2
- 239000008346 aqueous phase Substances 0.000 description 2
- 159000000007 calcium salts Chemical class 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 238000004090 dissolution Methods 0.000 description 2
- 239000012065 filter cake Substances 0.000 description 2
- 230000007935 neutral effect Effects 0.000 description 2
- 229920006395 saturated elastomer Polymers 0.000 description 2
- AZUYLZMQTIKGSC-UHFFFAOYSA-N 1-[6-[4-(5-chloro-6-methyl-1H-indazol-4-yl)-5-methyl-3-(1-methylindazol-5-yl)pyrazol-1-yl]-2-azaspiro[3.3]heptan-2-yl]prop-2-en-1-one Chemical compound ClC=1C(=C2C=NNC2=CC=1C)C=1C(=NN(C=1C)C1CC2(CN(C2)C(C=C)=O)C1)C=1C=C2C=NN(C2=CC=1)C AZUYLZMQTIKGSC-UHFFFAOYSA-N 0.000 description 1
- 102100022406 60S ribosomal protein L10a Human genes 0.000 description 1
- 101710155230 60S ribosomal protein L10a Proteins 0.000 description 1
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 1
- 239000002028 Biomass Substances 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 241000192125 Firmicutes Species 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 150000001298 alcohols Chemical class 0.000 description 1
- 150000001338 aliphatic hydrocarbons Chemical class 0.000 description 1
- 239000000908 ammonium hydroxide Substances 0.000 description 1
- 239000006286 aqueous extract Substances 0.000 description 1
- 239000012736 aqueous medium Substances 0.000 description 1
- 159000000009 barium salts Chemical class 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- BRPQOXSCLDDYGP-UHFFFAOYSA-N calcium oxide Chemical compound [O-2].[Ca+2] BRPQOXSCLDDYGP-UHFFFAOYSA-N 0.000 description 1
- 239000000292 calcium oxide Substances 0.000 description 1
- ODINCKMPIJJUCX-UHFFFAOYSA-N calcium oxide Inorganic materials [Ca]=O ODINCKMPIJJUCX-UHFFFAOYSA-N 0.000 description 1
- -1 calcium pseudomonate dihydrate Chemical class 0.000 description 1
- 125000004432 carbon atom Chemical group C* 0.000 description 1
- 239000000919 ceramic Substances 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 239000012043 crude product Substances 0.000 description 1
- 230000002939 deleterious effect Effects 0.000 description 1
- 238000011026 diafiltration Methods 0.000 description 1
- 239000003085 diluting agent Substances 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000009422 growth inhibiting effect Effects 0.000 description 1
- MPKNPWYCNYLGSC-UHFFFAOYSA-N heptane;4-methylpentan-2-one Chemical compound CCCCCCC.CC(C)CC(C)=O MPKNPWYCNYLGSC-UHFFFAOYSA-N 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 230000007062 hydrolysis Effects 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- GJRQTCIYDGXPES-UHFFFAOYSA-N iso-butyl acetate Chemical class CC(C)COC(C)=O GJRQTCIYDGXPES-UHFFFAOYSA-N 0.000 description 1
- FGKJLKRYENPLQH-UHFFFAOYSA-M isocaproate Chemical class CC(C)CCC([O-])=O FGKJLKRYENPLQH-UHFFFAOYSA-M 0.000 description 1
- OQAGVSWESNCJJT-UHFFFAOYSA-N isovaleric acid methyl ester Chemical class COC(=O)CC(C)C OQAGVSWESNCJJT-UHFFFAOYSA-N 0.000 description 1
- 238000000622 liquid--liquid extraction Methods 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- IBIKHMZPHNKTHM-RDTXWAMCSA-N merck compound 25 Chemical compound C1C[C@@H](C(O)=O)[C@H](O)CN1C(C1=C(F)C=CC=C11)=NN1C(=O)C1=C(Cl)C=CC=C1C1CC1 IBIKHMZPHNKTHM-RDTXWAMCSA-N 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 239000012466 permeate Substances 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- 239000003495 polar organic solvent Substances 0.000 description 1
- 238000012746 preparative thin layer chromatography Methods 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 235000017557 sodium bicarbonate Nutrition 0.000 description 1
- 229910000030 sodium bicarbonate Inorganic materials 0.000 description 1
- 239000012453 solvate Substances 0.000 description 1
- 238000000638 solvent extraction Methods 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 238000000108 ultra-filtration Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D407/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group C07D405/00
- C07D407/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group C07D405/00 containing two hetero rings
- C07D407/06—Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group C07D405/00 containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms
Definitions
- US patent no. 4222942 relates to a process for the isolation of mupirocin by extracting a solution of crude mupirocin preparation in a water immiscible organic solvent with an aqueous media, lowering the pH of said aqueous solution, and extracting the solution with a polar water immiscible organic solvent and thereafter adding a diluent which is sufficiently non-polar to reduce the polarity of, but is miscible with, said polar organic solvent so as to effect crystallization of mupirocin.
- the steps of this reaction scheme are manifold and, as a consequence, the scheme requires a large quantity of solvent.
- a cell free solution was obtained by centrifugation for 10 minutes in a laboratory centrifuge at 10,000 g (5° Celsius). About 90% of the mupirocin present in the fermentation broth was recovered in the final clarified solution.
- Example 5B Color removal by carbon treatment
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
- Plural Heterocyclic Compounds (AREA)
Abstract
Description
- The present invention relates to a method for purification of the antibiotic mupirocin (pseudomonic acid A).
- It is known that Pseudomonas fluorescens strains are able to biosynthesize, in addition to pseudomonic acid A, other related antibiotics designated by the letters B-D in small quantities [E. B. Chain, G. Mellows, J. Chem. Soc. Perkin Trans I. 318 (1977); J. P. Clayton et al., Tetrahedron Lett., 21, 881 (1980); P. J. O. Hanlon, N. H. Rogers, J. Chem. Soc. Perkin Trans I. 2665 (1983)]. Among the pseudomonic acid antibiotics, from a therapeutic point of view the most valuable is pseudomonic acid A, which has a growth inhibiting effect mainly against Gram positive bacteria.
- One method for the isolation of pseudomonic acid A from the antibiotic complex-containing culture broth is the liquid-liquid extraction. According to German Patent No.
2227739 andUS Patent No. 4289703 , soluble barium salts are added to the fermentation broth, then the microorganism cells with the insoluble inactive agents are separated by centrifugation and finally the antibiotics are extracted by methyl isobutyl ketone (MIBK). The antibiotics are then removed from the methyl isobutyl ketone extract by alkaline water and the resulting alkaline aqueous extract is cleaned by reextraction with methyl isobutyl ketone. The crude product obtained is chromatographed, and an ester derivative is prepared from the pseudomonic acid antibiotic complex and purified with preparative thin layer chromatography. The acid form of the pure antibiotic is obtained by hydrolysis. - Belgian Patent No.
870,855 -
US patent no. 4222942 relates to a process for the isolation of mupirocin by extracting a solution of crude mupirocin preparation in a water immiscible organic solvent with an aqueous media, lowering the pH of said aqueous solution, and extracting the solution with a polar water immiscible organic solvent and thereafter adding a diluent which is sufficiently non-polar to reduce the polarity of, but is miscible with, said polar organic solvent so as to effect crystallization of mupirocin. The steps of this reaction scheme are manifold and, as a consequence, the scheme requires a large quantity of solvent. -
US patent 6245921 discloses a process for the isolation of mupirocin comprising extracting an acidified culture broth using a chlorinated aliphatic hydrocarbon or isobutyl acetate, such that a mupirocin-containing extract is obtained; and purifying the mupirocin from said extract by distributing the extract between an aqueous phase and an organic phase comprising at least one organic solvent and evaporating the organic solvent. - None of the above methods are satisfactory from a commercial and ecological view. Accordingly, there remains a need for a novel method for purification of mupirocin, which is free from at least some of the disadvantages of the known processes and the application of which, in production scale, may result in a high yield of the recovery of the above-mentioned antibiotic.
- The present invention provides a method for purification of mupirocin, which meets at least some of the above defined objectives in that it is economically sound and, preferably, less ecologically deleterious, and results in a high yield of mupirocin having a high purity. The method comprises precipitation of mupirocin from an aqueous solution of mupirocin, and dissolution of the precipitate in an organic solvent.
- The present invention relates to a method for purifying mupirocin, comprising the steps of:
- a) causing or allowing precipitaton of mupirocin from an aqueous solution of mupirocin;
- b) recovering the precipitate;
- c) dissolving the recovered precipitate in an organic solvent; and optionally
- ci) recovering mupirocin from the solvent.
- The term "mupirocin" is meant to comprise mupirocin as defined in any of the above-mentioned references, as well as natural occurring and synthetic derivatives thereof. The term comprises the acidic form and salts thereof, such as the calcium salt, as well as solvates and polymorphic forms of these.
- In an embodiment of the invention, the solution of mupirocin is fermentation broth, such as clarified broth, such as clarified by centrifugation or filtration. The source of the fermentation broth is not crucial, and the broth can be obtained as disclosed in the prior art, for example as disclosed in the above-mentioned references, in
US 3977943 ,4071536 and4289703 ,WO 00/46389 WO 03/000910 - Mupirocin may be precipitated from the aqueous solution by adding an acid, such as an acid selected from the group consisting of acetic acid; citric acid; sulfuric acid; and hydrochloric acid. When using an acid, the pH during precipitation conveniently is in the range of 2.0 to 6.0, preferably in the range of 3.5 to 5.0, and most preferred in the range of 4.2 to 4.6. In one embodiment the solution is left stirring until mupirocin has precipitated, such as for a period of 10-40 minutes.
- The mupirocin containing precipitate may be recovered by centrifugation or filtration, centrifugation being preferred. The centrifugation should be carried out under such conditions that a gel is formed, e.g. by centrifuging for 5-30 minutes at 5000-15000g, or in an continuous centrifuge at 5000-15000g.
- In an embodiment of the invention, the recovered precipitate is dissolved in a polar, substantially water immiscible organic solvent. As solvent, a ketone, e.g. having the formula alkyl-C(=O)-alkyl, such as methyl isobutyl ketone; an ester, e.g. having the formula alkyl-C(=O)-O-alkyl, such as ethyl acetate; an alkanol, e.g. having the formula alkyl-OH, such as n-butanol; or a mixture comprising any of these can be used. Herein, the term "alkyl," when referring to alcohols, refers to aliphatic hydrocarbon groups of greater than four carbons and includes a straight or branched chain aliphatic hydrocarbon group that is saturated or unsaturated and, in other solvents, the term "alkyl" refers to a straight or branched chain aliphatic hydrocarbon group which is saturated or unsaturated and has 1, 2, 3, 4, 5, 6 or more carbon atoms.
- Presently it is preferred that the solvent is pure methyl isobutyl ketone (MIBK). That is the solvent should comprise more that 90% (such as more than 92%, more than 94%, more than 96%, or even more than 98% or 99%) MIBK. If needed, the mupirocin can be recovered from the solvent, such as by crystallization, optionally after water removal from the solvent.
- In another embodiment, the method of the invention comprises one or more further steps, in order to obtain a more pure antibiotic. Such steps might be applied before step a) and/or after step d), and/or between steps a) and b) and/or between steps b) and c) and/or after step c), and comprises steps selected from the group consisting of: a decolourisation step; a step for removal of hydrophilic components; a concentration step; an evaporation step; a clarification step; a water removal step; a drying step; a filtration step; an extraction step; and a crystallization step. It is presently preferred that the further step(s) are applied after the step d) above.
- The method of the invention is preferably carried out at a temperature between the freezing point of the solutions and room temperature, such as a temperature in the range 0-25°C, more preferred in the range 5-20°C. Optionally drying and evaporation steps can be carried out at an elevated temperature, such as in the range 20-50°C.
- The mupirocin prepared according to the invention is more pure than mupirocin obtained by the methods of the prior art. By the method of the invention, it is possible to obtain mupirocin having a purity of at least 93%, for example at least 95%, preferably least 96%, more preferably least 97% and most preferably even at least 98%. Typically, the mupirocin of the invention has a specific activity of at least 950 µg/mg, and preferably at least 960 µg/mg.
- Following the teaching of the prior art (e.g.
US 5569672 andUS 5594026 , the entire disclosure of which is incorporated by reference herein) it is also possible to generate calcium salts and different polymorphic forms of pseudomonic acid. - The use of the terms "a" and "an" and "the" and similar referents in the context of describing the invention (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. The terms "comprising", "having", "including" are to be construed as open-ended terms (i.e., meaning "including, but not limited to,") unless otherwise noted. Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein. All values mentioned herein should be understood as being preceded by "about", i.e. "7" should be understood as "about" 7. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or use of any and all examples, or exemplary language (e.g., "such as") provided herein intended merely to better illuminate the invention.
- Preferred embodiments of this invention are described herein, including the best mode known to the inventors for carrying out the invention.
- A fermentation broth containing mupirocin was obtained by fermenting a mupirocin producing culture of Pseudomonas fluorescens in a manner known per se, i.e. in line with the procedure of Example 1 in
US patent 3,977,943 . - 2 ℓ mupirocin containing whole culture fermentation broth from a laboratory fermentor was adjusted to pH = 7.1 (by addition of 1 M NaOH) and stirred at room temperature for 30 minutes. A cell free solution was obtained by centrifugation for 10 minutes in a laboratory centrifuge at 10,000 g (5° Celsius). About 90% of the mupirocin present in the fermentation broth was recovered in the final clarified solution.
- 5 ℓ mupirocin fermentation broth obtained as in example 1a was adjusted to pH = 8.3 (by addition of 3 M NaOH) and filtered through a 144 µm metal screen at room temperature, followed by ultra-filtration (Millipore Pellicon-2 Biomax-5). About 70 % of the mupirocin present in the fermentation broth was recovered in the final clarified solution.
- 2600 ℓ mupirocin fermentation broth obtained as in Example 1A was adjusted to pH = 7.5(by addition of 20% NaOH and filtered through a 0.1 µm ceramic filter (SCT/Exekia MembraloxP19-40,0.1-a-alu modules). Residual mupirocin was recovered by diafiltration of the biomass in batch mode with 3*500 ℓ water. Subsequently, the mupirocin containing permeate was concentrated by RO (Osmonics Desal DK membrane). It was found that 80% of the mupirocin present in the fermentation broth was recovered in the final clarified solution.
- 2 ℓ filtrate obtained as in example 1 c was transferred into centrifuge bottles and 40 mℓ concentrated acetic acid was slowly added under stirring. Resulting pH was 4.5. The sample was left stirring at room temperature for 30 minutes before centrifugation in a refrigerated centrifuge at 13,000 g for 20 minutes. The supernatant was poured off leaving a gel, containing 0.6 g mupirocin / g gel. Total recovery in the precipitation was 90%.
- 1000 ℓ clarified aqueous mupirocin solution obtained as in Example 1c was precipitated by lowering the pH to 4.5. About 50 ℓ 60 % acetic acid were mixed in line with the filtrate. The precipitate was collected by an industrial separator (Westfalia CSA-19), yielding a mupirocin gel containing 0.333 g mupirocin / g gel.
- 600 ml filtrate obtained as in Example 1A was transferred into centrifuge bottles and pH was adjusted to 4.4 by slowly adding 2 mt concentrated hydrochloric acid under stirring. The sample was left stirring at room temperature for 30 minutes before centrifugation in a refrigerated centrifuge at 13,000 g for 20 minutes. The supernatant was poured off leaving a gel.
- 3 mℓ aqueous precipitate obtained as in Example 2a was dissolved in 16 mℓ of 8% ammonium hydroxide. pH of the resulting clear solution (pH = 8.5) was adjusted to 4.4 by addition of 3 mℓ concentrated acetic acid. The sample was left at 4° Celsius over night before centrifugation in a refrigerated centrifuge at 13,000 g for 20 minutes. The supernatant was poured off leaving a gel. Total recovery in the second precipitation was 99%.
- Aqueous precipitate containing 25 g of mupirocin obtained as in Example 2a was dissolved in 250 mℓ of MIBK. The mixture was filtered.
- Aqueous precipitate containing 25 g of mupirocin obtained as in Example 2a was dissolved in 250 mℓ of ethyl acetate. The mixture was filtered.
- Example 5A: Removal of hydrophilic components 180 mℓ of mupirocin in MIBK obtained as in Example 4a was transferred to a separation funnel and extracted with 20 mℓ of MilliQ water. The two phases were left to separate and the aqueous phase was removed and discarded. The extraction of the organic phase was repeated 4 times and mupirocin was thereafter recovered from the organic phase.
- 180 mℓ of mupirocin in MIBK obtained as in Example 4a was added 4.5 g of activated carbon (Norit C extra) and left stirring for 30 minutes before the carbon was filtered off and the filter cake washed with 30 ml MIBK.
- 200mℓ of the organic solution obtained in Example 5a, containing 17 g mupirocin, was evaporated under vacuum In a laboratory rotary evaporator (60 mbar, 40°C). The evaporation was stopped when 102 mℓ solution was left in the product flask. 5 mℓ heptane was added and the solution was left with stirring at room temperature. After 22 hours with stirring the crystalline material was filtered. The filter cake was washed three times with 40 mℓ MIBK, and thereafter dried (< 50 mbar, 40°C). 13.2g dry product was recovered with a specific activity of 960 µg/mg (measured by HPLC).
- 100mℓ of organic solution, containing 17 g mupirocin, obtained as described in Example 5b, is added 150 mℓ heptane over about 2 hours with stirring. After about 24 hours with stirring the crystalline material is filtered, washed with MIBK/heptane (50/50) and dried in a vacuum drier (< 50 mbar, 40°C).
- 100mℓ of organic solution, containing 17 g mupirocin, obtained as described in Example 4b, is added 90 mℓ heptane with stirring. The solution is heated to 40°C, and held at this temperature for the rest of the procedure. The solution is seeded with crystalline product obtained from Example 6A. After three days with stirring the crystalline material is filtered, washed with MIBK/heptane (50/50) and dried in a vacuum drier (< 50 mbar, 40°C).
- Aqueous precipitate obtained as in Example 2A containing 25 g mupirocin is slowly dissolved with sodium hydroxide solution (1M, 45 mℓ) to give a neutral solution (pH 7). The solution is filtered. Calcium chloride (6.3 g) is added the mixture and stirring is maintained to give a clear solution. After standing for 20 hours the crystalline product is filtered off, washed with water (50 mℓ) and dried (50 mbar, 40°C) to give calcium pseudomonate dihydrate.
- Aqueous precipitate obtained as in Example 2A containing 25 g mupirocin is added 50% aqueous methanol to a volume of 250 mℓ. Calcium oxide (1.8 g) is added to give a neutral solution (pH 7). The solution is evaporated in order to remove methanol. The methanol-free residue (120 mℓ) is diluted with water (50 mℓ) and allows crystallizing with stirring in room temperature. After 24 hours the mixture is filtered and the collected product is washed by slurrying with water (50 mℓ). The crystalline product is dried in a vacuum oven (50 mbar, 40°C).
- Quality of process and final product:
US 4,222,942 Ex1US 4,222,942 Ex2Invention Ex. 6a Purity (final product) 92-93 % 96% Yield (from filtrate to final product) 31 % 47% 47% Solvent use Liter/g product 2.5 1.7 0.03 "Steps" 8 9 3 "Steps" = number of steps required to obtain a concentrated solvent extract from a clarified broth / filtrate
Claims (11)
- A method for purifying mupirocin, comprising the steps of:a) causing or allowing precipitation of mupirocin from an aqueous solution of mupirocin, wherein the pH of the solution during precipitation is in the range of 2.0 to 6.0;b) recovering the precipitate;c) dissolving the recovered precipitate in an organic solvent; and optionallyd) recovering mupirocin from the solvent.
- A method according to Claim 1, characterized in that mupirocin is precipitated by adding an acid, such as an acid selected from the group consisting of acetic acid; citric acid: sulfuric acid; and hydrochloric acid.
- A method according to Claim 1 or 2, characterized in that the pH of the solution during precipitation is in the range of 3.5 to 5.0.
- A method according to Claim 1 or 2, characterized in that the pH of the solution during precipitation is in the range of 4.2 to 4.6.
- A method according to any of the claim 1 to 4, characterized in that the solution of mupirocin is clarified fermentation broth.
- A method according to any of the preceding claims, wherein the mupirocin containing precipitate is recovered in step b) by centrifugation or filtration.
- A method according to any of the preceding claims, wherein, in step c), the organic recovered precipitate is dissolved in a polar, water immiscible or moderately soluble organic solvent.
- A method according to any of the preceding claims, wherein the organic solvent is a ketone, ; an ester, ; or an alkanol, ; or a mixture comprising any of these.
- A method according to any of the preceding claims, wherein the organic solvent is pure methyl isobutyl ketone.
- A method according to any of the preceding claims, wherein mupirocin is recovered from the solvent by crystallization.
- The method according to any of the preceding claims, further comprising one or more additional steps selected from the group consisting of; a decolourisation step; a step for removal of hydrophilic components; a concentration step; an evaporation step; a clarification step; a water removal step; a drying step; a filtration step; an extraction step; and a crystallization step.
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
DKPA200500261 | 2005-02-21 | ||
PCT/EP2006/001672 WO2006087237A1 (en) | 2005-02-21 | 2006-02-21 | Purification of mupirocin |
Publications (2)
Publication Number | Publication Date |
---|---|
EP1856111A1 EP1856111A1 (en) | 2007-11-21 |
EP1856111B1 true EP1856111B1 (en) | 2014-01-29 |
Family
ID=36609364
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
EP06723096.1A Active EP1856111B1 (en) | 2005-02-21 | 2006-02-21 | Purification of mupirocin |
Country Status (8)
Country | Link |
---|---|
US (1) | US7619102B2 (en) |
EP (1) | EP1856111B1 (en) |
JP (1) | JP5101305B2 (en) |
CN (1) | CN101124221B (en) |
AU (1) | AU2006215708B2 (en) |
CA (1) | CA2598565C (en) |
ES (1) | ES2455518T3 (en) |
WO (1) | WO2006087237A1 (en) |
Families Citing this family (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
GB2441328A (en) * | 2006-08-30 | 2008-03-05 | Alpharma Aps | A method for obtaining mupirocin calcium |
CN101591333B (en) * | 2009-07-02 | 2012-01-04 | 山东健威生物工程有限公司 | Method for purifying pseudomonas acid A |
CN104370896A (en) * | 2013-11-29 | 2015-02-25 | 江苏汉邦科技有限公司 | Pseudomonas elodea rhzomorph purifying method |
CN109053707B (en) * | 2018-09-19 | 2021-08-17 | 福建康鸿生物科技有限公司 | Purification method of mupirocin |
TW202214611A (en) | 2020-08-25 | 2022-04-16 | 大陸商杭州中美華東製藥有限公司 | Method for extracting mupirocin |
Family Cites Families (9)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
GB1395907A (en) | 1971-06-12 | 1975-05-29 | Beecham Group Ltd | Antibiotics |
US4071536A (en) | 1971-06-12 | 1978-01-31 | Beecham Group Limited | Antibiotics |
US3977943A (en) * | 1971-06-12 | 1976-08-31 | Beecham Group Limited | Antibiotics |
CA1103264A (en) * | 1977-09-30 | 1981-06-16 | Norman H. Rogers | Purification of pseudomonic acid |
GB8415579D0 (en) | 1984-06-19 | 1984-07-25 | Beecham Group Plc | Compounds |
US5594026A (en) | 1990-12-11 | 1997-01-14 | Smithkline Beecham Group P.L.C. | Polymorphs of crystalline mupirocin |
TR200102234T2 (en) * | 1999-02-03 | 2002-10-21 | Biogal Gyogyszergyar Rt. | Process for the preparation of pseudomonic acid A antibiotic by microbiological method |
HUP0105286A3 (en) * | 1999-02-03 | 2003-03-28 | Biogal Gyogyszergyar | Process for the isolation of pseudomonic acid a from pseudomonic acid complex-containing culture broth |
CA2448547A1 (en) | 2001-06-21 | 2003-01-03 | Biogal Gyogyszergyar Rt. | Metabolic controlled fermentation process for pseudomonic acid production |
-
2006
- 2006-02-21 CN CN2006800055544A patent/CN101124221B/en active Active
- 2006-02-21 US US11/816,439 patent/US7619102B2/en active Active
- 2006-02-21 AU AU2006215708A patent/AU2006215708B2/en active Active
- 2006-02-21 CA CA2598565A patent/CA2598565C/en active Active
- 2006-02-21 JP JP2007555552A patent/JP5101305B2/en active Active
- 2006-02-21 WO PCT/EP2006/001672 patent/WO2006087237A1/en active Application Filing
- 2006-02-21 ES ES06723096.1T patent/ES2455518T3/en active Active
- 2006-02-21 EP EP06723096.1A patent/EP1856111B1/en active Active
Also Published As
Publication number | Publication date |
---|---|
US7619102B2 (en) | 2009-11-17 |
ES2455518T3 (en) | 2014-04-15 |
AU2006215708B2 (en) | 2011-01-27 |
CA2598565C (en) | 2013-06-25 |
US20080234503A1 (en) | 2008-09-25 |
CA2598565A1 (en) | 2006-08-24 |
JP5101305B2 (en) | 2012-12-19 |
AU2006215708A1 (en) | 2006-08-24 |
JP2008530168A (en) | 2008-08-07 |
EP1856111A1 (en) | 2007-11-21 |
CN101124221A (en) | 2008-02-13 |
CN101124221B (en) | 2012-05-23 |
WO2006087237A1 (en) | 2006-08-24 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
FI67222B (en) | FOERFARANDE FOER RENING AV KLAVULANSYRA OCH SAOSOM MELLANPROTUKT ANVAENDBARA AMINSALT AV KLAVULANSYRA SAMT FOERFARANDE FOER FRAMSTAELLNING AV DESSA | |
RU2081121C1 (en) | Method of synthesis of clavulanic acid or its pharmaceutically acceptable salts and ethers, clavulanic acid salt with amine | |
EP1856111B1 (en) | Purification of mupirocin | |
US4222942A (en) | Isolation of organic acids | |
EP0005614B1 (en) | Lithium pseudomonate, process for its isolation and its hydrolysis | |
AU771214B2 (en) | Process for the isolation of pseudomonic acid a from pseudomonic acid complex-containing culture broth | |
US3127315A (en) | Hypocholesterolemic agent m- | |
EP0941229B1 (en) | Purification of fermented clavulanic acid | |
JP3207870B2 (en) | Cyclic depsipeptide and method for producing the same | |
JP2873894B2 (en) | Cyclic depsipeptide and method for producing the same | |
CN1312814A (en) | Improved process for preparing salts and esters of clavulanic acid | |
JP2001521393A (en) | Method for isolating pharmaceutically acceptable alkali metal salts of clavulanic acid | |
RU2515936C2 (en) | Highly pure pentamycin | |
JP2512050B2 (en) | Novel antifungal antibiotic dexylosylvenanomycin B and method for producing the same | |
EA017121B1 (en) | Process for the preparation of a potassium salt of penicillin g | |
US20100055749A1 (en) | Process for the production of clavulanic acid | |
JPH0665277A (en) | Antibiotic dihydroaldecalmycin and its production |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PUAI | Public reference made under article 153(3) epc to a published international application that has entered the european phase |
Free format text: ORIGINAL CODE: 0009012 |
|
17P | Request for examination filed |
Effective date: 20070730 |
|
AK | Designated contracting states |
Kind code of ref document: A1 Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HU IE IS IT LI LT LU LV MC NL PL PT RO SE SI SK TR |
|
DAX | Request for extension of the european patent (deleted) | ||
RAP1 | Party data changed (applicant data changed or rights of an application transferred) |
Owner name: AXELLIA PHARMACEUTICALS APS |
|
RAP1 | Party data changed (applicant data changed or rights of an application transferred) |
Owner name: XELLIA PHARMACEUTICALS APS |
|
RAP1 | Party data changed (applicant data changed or rights of an application transferred) |
Owner name: SYNGEN BIOTECH CO., LTD. |
|
17Q | First examination report despatched |
Effective date: 20120525 |
|
GRAP | Despatch of communication of intention to grant a patent |
Free format text: ORIGINAL CODE: EPIDOSNIGR1 |
|
INTG | Intention to grant announced |
Effective date: 20130828 |
|
GRAS | Grant fee paid |
Free format text: ORIGINAL CODE: EPIDOSNIGR3 |
|
GRAA | (expected) grant |
Free format text: ORIGINAL CODE: 0009210 |
|
AK | Designated contracting states |
Kind code of ref document: B1 Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HU IE IS IT LI LT LU LV MC NL PL PT RO SE SI SK TR |
|
REG | Reference to a national code |
Ref country code: GB Ref legal event code: FG4D |
|
REG | Reference to a national code |
Ref country code: CH Ref legal event code: EP |
|
REG | Reference to a national code |
Ref country code: AT Ref legal event code: REF Ref document number: 651492 Country of ref document: AT Kind code of ref document: T Effective date: 20140215 |
|
REG | Reference to a national code |
Ref country code: IE Ref legal event code: FG4D |
|
REG | Reference to a national code |
Ref country code: DE Ref legal event code: R096 Ref document number: 602006040233 Country of ref document: DE Effective date: 20140313 |
|
REG | Reference to a national code |
Ref country code: ES Ref legal event code: FG2A Ref document number: 2455518 Country of ref document: ES Kind code of ref document: T3 Effective date: 20140415 |
|
REG | Reference to a national code |
Ref country code: AT Ref legal event code: MK05 Ref document number: 651492 Country of ref document: AT Kind code of ref document: T Effective date: 20140129 |
|
REG | Reference to a national code |
Ref country code: NL Ref legal event code: VDEP Effective date: 20140129 |
|
REG | Reference to a national code |
Ref country code: LT Ref legal event code: MG4D |
|
PG25 | Lapsed in a contracting state [announced via postgrant information from national office to epo] |
Ref country code: IS Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140529 Ref country code: LT Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 |
|
PG25 | Lapsed in a contracting state [announced via postgrant information from national office to epo] |
Ref country code: FI Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: CY Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: SE Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: NL Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: PT Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140529 Ref country code: AT Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 |
|
PG25 | Lapsed in a contracting state [announced via postgrant information from national office to epo] |
Ref country code: LV Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: BE Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 |
|
REG | Reference to a national code |
Ref country code: DE Ref legal event code: R097 Ref document number: 602006040233 Country of ref document: DE |
|
PG25 | Lapsed in a contracting state [announced via postgrant information from national office to epo] |
Ref country code: EE Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: RO Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: CZ Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: DK Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 |
|
PG25 | Lapsed in a contracting state [announced via postgrant information from national office to epo] |
Ref country code: PL Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: SK Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 |
|
REG | Reference to a national code |
Ref country code: IE Ref legal event code: MM4A |
|
PLBE | No opposition filed within time limit |
Free format text: ORIGINAL CODE: 0009261 |
|
STAA | Information on the status of an ep patent application or granted ep patent |
Free format text: STATUS: NO OPPOSITION FILED WITHIN TIME LIMIT |
|
26N | No opposition filed |
Effective date: 20141030 |
|
PG25 | Lapsed in a contracting state [announced via postgrant information from national office to epo] |
Ref country code: IE Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES Effective date: 20140221 |
|
REG | Reference to a national code |
Ref country code: DE Ref legal event code: R097 Ref document number: 602006040233 Country of ref document: DE Effective date: 20141030 |
|
PG25 | Lapsed in a contracting state [announced via postgrant information from national office to epo] |
Ref country code: SI Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 |
|
REG | Reference to a national code |
Ref country code: FR Ref legal event code: PLFP Year of fee payment: 11 |
|
PG25 | Lapsed in a contracting state [announced via postgrant information from national office to epo] |
Ref country code: MC Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: BG Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 |
|
PG25 | Lapsed in a contracting state [announced via postgrant information from national office to epo] |
Ref country code: GR Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140430 Ref country code: IT Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 |
|
PG25 | Lapsed in a contracting state [announced via postgrant information from national office to epo] |
Ref country code: LU Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES Effective date: 20140221 Ref country code: TR Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT Effective date: 20140129 Ref country code: HU Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT; INVALID AB INITIO Effective date: 20060221 |
|
REG | Reference to a national code |
Ref country code: FR Ref legal event code: PLFP Year of fee payment: 12 |
|
REG | Reference to a national code |
Ref country code: FR Ref legal event code: PLFP Year of fee payment: 13 |
|
P01 | Opt-out of the competence of the unified patent court (upc) registered |
Effective date: 20230607 |
|
PGFP | Annual fee paid to national office [announced via postgrant information from national office to epo] |
Ref country code: GB Payment date: 20231208 Year of fee payment: 19 |
|
PGFP | Annual fee paid to national office [announced via postgrant information from national office to epo] |
Ref country code: ES Payment date: 20240305 Year of fee payment: 19 |
|
PGFP | Annual fee paid to national office [announced via postgrant information from national office to epo] |
Ref country code: DE Payment date: 20231218 Year of fee payment: 19 Ref country code: CH Payment date: 20240301 Year of fee payment: 19 |
|
PGFP | Annual fee paid to national office [announced via postgrant information from national office to epo] |
Ref country code: FR Payment date: 20240129 Year of fee payment: 19 |